The objective of this proposal is to understand the synthesis, importance and regulation of the phospholipid component of biological membranes through a combined enzymological and genetic approach in the bacterium, Escherichia coli, which is a suitable model system for studying such fundamental processes. An understanding of the cellular control mechanisms which operate to regulate the quantity and quality of phospholipid synthesized will be sought by the isolation of regulatory mutants. The importance of membrane phospholipid composition to the function of membranous enzymes will be investigated by obtaining membranes of altered composition from mutants, from cells by the in vivo action of chemical agents or in vitro by the action of certain phospholipases or by exchange of phospholipids. Phospholipid synthesis and membrane assembly will be studied by isolating conditional mutants defective in phospholipid synthesis by H3-oleic acid and H3-glycerol suicide techniques and by a localized mutagenesis procedure. The availability of such mutants should allow a detailed understanding of the phospholipid biosynthetic pathway, the functional significance of each phospholipid class, a better understanding of membrane assembly, and the coordination of DNA, RNA, protein and cell wall synthesis with phospholipid synthesis and membrane assembly. Studies on the basic involvement of phospholipids in membrane processes in the E. coli model system, are particularly promising since they can draw on the extensive physiological, biochemical and genetic studies on this organism, and should provide direction for investigations into mammalian cells. An understanding of the contribution of membrane phospholipid to the structure and function of the membrane should contribute substantially to understanding important biological processes and understanding certain disease states.